Control and track axonal growth and axonal pathfinding over time
The development of the nervous system is one of the most fascinating and at the same time least understood biological processes. FluidFM technology provides a new tool in studying axonal guidance and axonal outgrowth for in-vitro research at the single-neuron level. By combining unique nanoprinting, injection, extraction, and single cell placing technologies, FluidFM technology allows to:
Create customized patterns to study axonal guidance and outgrowth forming neuronal circuits
Precisely and locally dispense or inject neuromodulators or other soluble substances to study intra-cellular axonal trafficking
Manipulate single neurons to establish in vitro disease models for neurological disorders
Study axonal guidance and outgrowth with FluidFM
FluidFM enables creating customized patterns of, for example, attractive and repellent signals and hence the formation of neuronal circuits in vitro in a controlled manner. Also, using its injection feature, individual neurons can be transfected which opens new possibilities in studying individual genes and proteins in a single or multicellular context.
Unfold to learn more about axonal pathfinding
The axonal pathfinding is steered by short- and long-range axon guidance cues (like netrins, semaphorins or neuronal cell adhesion molecule (NrCAM)) as well as attractive or repulsive guidance information that enable axonal navigation and help the axonal growth cone decide in which direction and along which choice points it has to grow or whether it has to stop growing and form synapses. Signals, such as chemotactic signals, must be received by guidance receptors (e.g. Semaphorin/Plexin or Robo/Slit) and properly interpreted by the growing neuron to control axonal outgrowth. While many questions remain open, research of the last decades have revealed a number of important and conserved genes that play a role in axon guidance.
Image shows PLL line in green, printed with FluidFM between two groups of neurons. In red, neurite growth driven by PLL can be seen. Image courtesy of Harald Dermutz, ETH Zurich, Switzerland.
The series of image above shows HeLa-GFP cells before, during, and after extraction of cytosolic content by a FluidFM Nanosyringe. Image courtesy of Orane Guillaume-Gentil, ETH Zurich, Switzerland.
Study intra-cellular axonal trafficking with FluidFM
With FluidFM soluble molecules can be administered on or into the cell at any location of the cell, even directly into the nucleus. Hence local dispensing of neuromodulators at the growth cone or anywhere at the axon is possible. Likewise, FluidFM also allows to extract material from the cell while keeping the cell viable e.g. for further observation or a consecutive extraction. This is particularly interesting for time-consecutive proteome or transcriptome analysis or single cell sequencing.
Unfold to learn more about axonal trafficking
Neurons are composed of a cell body where the nucleus is located, of several branched dendrites that receive signals from other neurons and of one lengthy single axon that sends signals to other cells. The lengthy structure of axons poses specific challenges in intra-cellular trafficking of vesicles or other cargos but also local axonal translation of proteins. Here mRNA must be transported along the entire axon while its translation is suppressed. This fascinating mechanism however is still not well understood and its study so far encompasses complex culture systems with compartmentalized chambers (like Campenot chamber) and microfluidics. New tools to improve studying local processes is therefore highly needed, also to open opportunities to apply state of the art transcriptomics or proteomics.
Studying neurological disorders with FluidFM
FluidFM provides a new comprehensive tool to create in vitro disease models for diseases and research in the neuroscience field. By its single cell manipulation technology, it is possible to transfect or even genetically engineer single neurons or other hard-to-transfect cells. Furthermore, by its nanoprinting and single cell pick-and-place features it allows placing specific individual cells, even cells from different wells, next to each other to study cell-cell interactions and communication or creating neuronal circuits between specific cells. And finally, one can also study the characteristics of single cells upon mechanical or chemical stimulation by dispensing or even cytosolic or nuclear injection of neuromodulators or any other soluble compounds.
Unfold to learn more about neurological disorders
Axon guidance and loss of neuronal circuits is affected by or implicated in many disorders of the nervous system. Neurodevelopmental disorders, like schizophrenia or autism spectrum disorders, are all linked to aberrant development of neural circuits. In neurodegenerative disorders such as Alzheimer’s or Parkinson’s disease, the disintegration of neural circuits plays a key role. Spinal cord injuries, for example caused by a trauma, are correlated with loss of neural circuits and the inability of spinal cord central nerve cells to regenerate, in contrast to peripheral nerves that can regenerate and reinnervate tissue. In Multiple Sclerosis, the loss of oligodendrocytes that form a myelin sheath along the axons to help neurons to carry electrical signals, leads to a breakdown of axons and eventually the formation of scar-like plaques. Not only because of MS, which is believed to be caused by an autoimmune response, the field of neuroimmunology has raised a lot of attention in the recent years. In summary, an understanding of axon guidance is key not only to comprehend neuronal development and how neural circuits and hence the brain work but also to increase our knowledge in finding therapies against devastating diseases.
Neuron expressing GFP 24h after injection of a plasmid encoding GFP using FluidFM. Image courtesy of Sen Yan, Jinan University, Guangzhou, China.